6 alpha-fluoro-16 alpha-hydroxy steroids



United States Patent 6a-FLUORO-16u-HYDROXY STEROIDS Barney J. Magerlein, Kalamazoo, William P. Schneider, Kalamazoo Township, Kalamazoo County, and Oldrich K. Sebek and George B. Spero, Kalamazoo, Mich., assignors to The Upjohn Company, Kalamazoo, Mich a corporation of Michigan No Drawing. Application February 19, 1958 Serial No. 716,004

19 Claims. (Cl. 260--397.45)

.The present invention relates to steroid compounds and is more particularly concerned with 6a-fluoro-11B, 16a,l.7a trihydroxy 4-pregnene3,20-dione, 6a,9u-difluoro 11fi,16o ,17ot trihydroxy-4-pregnene-3,20-dione, 6a-

wherein the 1,2-carbon atom linkage is selected from the linkages consisting of single bond and double bond linkages, Y is selected from the group consisting of hydrogen and fluorine, X is selected from the group consisting of the carbonyl radical C=O) and the B-hydroxymethylene radical and R is selected from the group consisting of hydrogen and the acyl radical of an organic carboxylic acid, preferably a hydrocarbon carboxylic acid containing from one to twelve carbon atoms, inclusive.

The newcompounds 6a-fluoro-11p,16u,17a-trihydroxy- 4-pregnene-3,20-dione, 6a,9ot-difluoro-l1B,l6a,17a-trihydroxy 4-pregnene-3,20-dione, 6a-fluoro-l1,8,l6a,17a-trihydroxy 1,4 pregnadiene 3,20 1' dione, 6oc,9oc-difluor0- 115,16,17ot-trihydroxy-1,4 pregnadiene 3,20-dione, their l6-esters and the ll-keto analogues and esters thereof, are highly active adrenocortical hormones having greater glucocorticoid and anti-inflammatory activity than hydrocortisone or cortisone. In addition these compounds have diuretic activity and have salt-losing properties which make them especially well suited in the management of chronic congestive heart failure and in the treatment of cirrhosis of the liver, the nephrotic and adrenogenital syndromes and the treatment of eclampsia and preeclampsia.

Patented June 1%, 35E558 trihydroxy 1,4 pregnadiene 3,20 dione, the l6-esters thereof, the ll-keto analogues and the l6-esters thereof can be given in oral, parenteral or topical compositions. The compounds can be administered to the animal organism in conventional dosage forms such as pills, tablets and capsules for oral use or in conventional liquid forms as are used with natural and synthetic cortical steroid hormones for injection use. For topical use they can be administered in the form of ointments, creams, lotions and the like with or without co-acting antibiotics, germicides and the like.

The processof the present invention comprises microbiological hydroxylation of 6a-fiuoro-11,8,17a-dihydroxy- 4-pregnene-3,20-dione, 60,9ot-difillOIO-115,170c-dihYd10XY- 4-pregnene-3,20-dione, and the A -analogues thereof to produce the corresponding 16 -hydroxy compounds. Esterification of the thus produced 16-hydroxylated compounds is productive of the l6-esters. The corresponding ll-keto analogues of these 16-hydroxylated compounds are obtained by oxidation of the 11 -hydroxyl group of the above l6-esterified compounds with an oxidation agent such as chromic acid. If the free alcohols of the ll-keto compounds thus produced are desired, the additional step of hydrolyzing the 16-esters such as with an alkali metal base is necessary.

Starting materials for the present invention are 60:- fluoro llfl,l7a-dihydroxy-4-pregnene-3,20-dione, 6a,9izdifluoro 11,43,170: dihydroxy-4-pregnene-3,ZO-dione, 6afluoro 11,3,17ot dihydroxy 1,4 pregnadiene 3,20 dione, and 6ot,9 x-diiluoro-llfi,17a-dihydroxy-1,4-pregnadiene-3,20-dione, prepared as shown in the preparations.

In the bioconversion step ofthe present invention, the

operational conditions and reaction procedure and details can be those already known in the art of steroid bioconversion as illustrated by the Murray et a1. U. S. Patent 2,602,769 issued July 8, 1952, utilizing however the action of an organism of the genus Streptomyces. Among the species which are useful in the fermentation step of the present invention are Streptomyces roseochromogenus (Waksman collection 3689,), Streptomyces sp. (A. T. C. C. 11009), and Streptomyces roseochromo genus (A. T. C. C. 3347). The selected species of actinomycete is grown on a medium suitably containing assimilable non-steroid carbon, illustratively carbohydrates, such as dextrose; assimilable nitrogen, illustratively soluble or insoluble proteins, peptones or amino acids; and mineral constituents, illustratively sodium or ammonium phosphate and magnesium sulfate. The medium may desirably have a pH before innoculation of between about 6.5 to about 7.8 though a higher or lower pH may be used. A pH of between about 6.8 and about 7.4 is preferred for the growth of actinomycetes and a temperature range from about 20 to about 35 degrees centigrade with about 20 to 32 degrees centigrade preferred.

The growth period required before the steroid to be fermented is exposed to the actinomycete does not appear to be critical, for example, the steroid may be added either before sterilization of the medium, at the time of innoculation the medium or at some time later, for example, 24 ,or' 48 hours later. The addition of steroid substrate to be fermented may be accomplished in any suitable manner, such as by dispersing the steroid substrate, either alone with a dispersing agent, or in solution in an organic solvent. Either submerged or surface culture procedures may be used with facility, although submerged culture is preferred.

The temperature during the period of fermentation of the steroid may be the same as that found suitable for the growth of the organism. It need be maintained 3 only within such range as supports life, active growth, or the enzyme activity of the streptomycete.

The time required for the fermentation of steroid varies somewhat with the procedure. When the steroid is added to the actinomycete after substantial growth of the organism, for example, after 16 to- 24 hours at optimum temperature, the conversion of steroid substrate begins immediately and is substantially complete in from two to ten days, five days being generally satisfactory.

After completion of the steroid fermentatiomthe resulting transformed steroid is recovered from he fermentation reaction mixture by extracting the fermentation reaction mixture, including the fermentation liquid and mycelium with an organic solvent for steroids, for example, methyl isopropyl ketone, methylene chloride, chloroform, carbon tetrachloride, ethylene chloride, trichloroethylene, ether, .amyl acetate, benzene, and the like. The fermentation liquor and mycelium can be separated and then separately extracted with suitable solvents. The extracts can be combined, either before or after washing with an alkaline solution, illustratively sodium bicarbonate, suitably dried, a for example, over 7 anhydrous sodium sulfate, and the resulting purified transformed steroid obtained by recrystallization from organic solvents, by trituration or by chromatography in order to isolate the thus obtained steroids from the other transformation products. Bioconversion of 6tz-fiuoro-11 3,17otdihydroxy-4-pregnene-3,20-dione, 606,90t-dlfill0101 113,170:- dihydroxy-4-pregene-3,ZO-dione, 6Ct-fi11OI'O-11/8,170t dihy droxy-1,4-pregnadiene-3,20-dione, and 60,90c-Cliflll010- 11/8,l7oc-dihydroxy-l,4-pregnadiene-3,20 dione according to the fermentation procedure above described, is productive of fiix-fiuoro-l1p,16a,17a-trihydroxy-4-pregnene-3,20 dione, 6a, 9oi-difit10rO-l 1,8,16a,17a-trihydrOXyA-pregnene- 3,20-dione, 605- fiuoro-l15,16a,17ot-trihydroxy-1,4-pregnadiene-3,20-dione, and 6m,9o.-difiuoro-1lfi,16a,17ot-trihydroxy l,4-pregnadiene-3,20-dione, respectively.

The 16-hydroxylated compounds thus produced can be esterified to produce the corresponding 16-esters. This reaction can be performed under esterification conditions known in the art, e. g., by the reaction of the hydroxy compound with the selected acid halide, e. g., acid chloride or acid bromide, the anhydride of a hydrocarbon carboxylic acid, or by'reaction with the selected acid, in the presence of an 'esterification catalyst or with an ester under ester exchange reaction conditions. Reaction conditions which are apt to affect the labile llfi-hydroxy group or 6-fiuoro group should be avoided. Compounds thus produced include the 16-acyloxy compounds represented by Formula I wherein X is fl-hydroxymethylene radical and wherein R is the acyl radical of an organic carboxylic acid, preferably a hydrocarbon carboxylic acid containing from one to twelve carbon atoms, inclusive, e. g., formic, propionic, butyric, isobutyric, valeric, isovaleric, trimethylacetic, Z-methylbutyric, 3- ethylbutyric, hexanoic, diethylacetic, triethylacetic, heptanoic, octanoic, a-ethylisovaleric, succinic, a cyclic acid, e. g., cyclopropylideneacetic, cyclopentylformic, cyclopentylacetic, B-cyclohexylpropionic, cyclohexylformic, cyclohexylacetic, an aryl or alkaryl acid, e. g., benzoic, 2- 3- or 4-methylbenzoic, 2,3-, 2,4-, 2,5-, 2,6-, 3,4- and 3,5- dimethylbenzoic, ethylbenzoic, 2,4,6-trimethylbenzoic, 2,4,6-triethylbenzoic, u-naphthoic, 3-methyl-a-naphthoic, an aralkyl acid, e. g., phenylacetic, phenylpropionic, diphenylacetic, triphenylacetic, an unsaturated acid, e. g., acrylic, maleic, vinyl .acetic, propiolic, undecolic, etc. Illustrative of the esters thus produced are the 16-acylates such as 6ot-fluoro-11,8,16ot,l7ot-trihydroxy-1,4-pregnadiene- 3,20-dione l6-acetate, 6ll'r'fiUOIO-11,160L,170CtrlhydrOXy- 1,4-pregnadiene-3,ZO-dione l6 acetate, 6a, 9a-difluoro-11B, 16a,17ot-trihydroxy-4-pregnene-3,ZO-dione 16-acetate, 64x, 9a-difluoro-11,8,16a,17a-trihydroxy-1,4-pregnadiene-3,20 dione 16-acetate, and the like. 7

The llfi-hydroxyl of the thusproduced l6-acylates can be oxidized to the correspondingll-ketone with an oxidizing agent. Oxidizing agents such as chromic acid, potassium dichromate, a halo-amide, and the like are operative. The oxidation can be carried out by a variety of methods, such as, for example, by oxidizing the said llfi-hydroxy steroid in acetic acid-water solution with chromium trioxide, using molar quantities or a slight excess, such as from ten to thirty percent excess, or by oxidizing with a haloamide or imide of an acid, such as N-bromoacetamide, N-chlorosuccinimide, or N-bromosuccinimide dissolved in pyridine, dioxane, or other suitable solvents. At the conclusion of the desired oxidation reaction, the excess oxidant is generally destroyed by addition of methyl alcohol, ethyl alcohol, and the like for the chromic acid oxidant or a bisulfite for N-bromoacetamide, N-bromosuccinimide and other N-haloacylamides and irnides. Thereafter, the resulting ll-keto product is recovered by conventional means, such as by dilution with water andextraction with a water-immiscible solvent, e. g., methylene chloride, ether, benzene, toluene, ethyl acetate, or the like. Illustrative of the ll-keto l6- acylates thus produced are, for example, 60c-flLIO1O-l6oz, 17u-dihydroxy-4-pregnene-3,11,20-trione 16-acetate, 60c, 9a-difluoro-16ot,17a-dihydroxy-4-pregnene-3,11,20 trione 16-acetate, dot-fiuoro16a,17a-dihydroxy-1,4-pregnadiene 3,11,20-trione l6-acetate, 611,911 difiuoro :,17ot dihydroxy-l,4-pregnadiene-3,11,20-trione 16-acetate, and the like.

The ll-keto 16-acylates thus produced may, if desired, be converted to the free alcohols, i. e., 6a-fluoro-16a,17adihydroxy-4-pregnene-3,l1,20-trione, 60:,90: (llfillOrO-l6oc, 17a-dihydroxy-4-pregnene-3,11,20-trione, 6c: fiu0rO-l6oz, l7u-dihydroxy-l,4-pregna-diene-3,11,20-trione and 611,90:- difiuoro-16a,17a-dihydroxy 1,4 pregnadiene-3,1l,20- trione. The alcohols are obtained from the acylates by hydrolysis in accordance with general hydrolysis procedures known in the art. A preferred procedure is to employ at least a molar equivalent of an alkali-metal bicarbonate in a substantially oxygen-free solution of a mixture of a lower alkanol and water. The hydrolysis reaction is carried out at a temperature between ten and thirty degrees centigrade while protecting the mixture from at mospheric oxygen. After the hydrolysis is complete, the reaction mixture is neutralized with an acid, e. g., acetic acid, and the hydrolyzed product recovered from the reaction mixture by evaporation and .crystallization, extraction with methylene chloride, or the like. The IS-esters when desired, can again be prepared by esterification of the hydroxyls by esterification procedures hereinbefore described. The preferred esters are those derived from an organic carboxylic acid, preferably a hydrocarbon carboxylic acid containing from one to twelve carbon atoms, inclusive.

The following preparations and examples are illustrative of the process and products of the present invention but are not to be construed as limiting.

PREPARATION 1 The 3-ethylene ketal of methyl 3,11-diketo-5a-6ot- Oxl'do-I7[20]-[cis]-pregnene-21-0ate To a solution of 5.0 grams of the 3-ethylene ketal of methyl 3,11 diketo 4,17 (20)-[cis]-pregnadien-21-oate, prepared in the manner described in U. S. Patent 2,707,- 184, in 100 milliliters of chloroform was added a chilled solution of 1.9 grams of perbenzoic acid dissolved in 31.5 milliliters of chloroform. The solution was maintained at about four degrees centigrade for 24 hours, and then at room temperature for 72 hours. The solution was then washed with a five percent aqueous solution of sodium bicarbonate and then with water. The chloroform layer was separated, dried and the solvent distilled to give a residue of 5.3 grams of solid. Crystallization of this solid from methanol gave 2.24 grams of product melting at to degrees centigrade and after two crystallizations from methanol, there was obtained pure ,3-ethylene ketal of methyl 3,1l-.diket05a,6a-0Xid()- l7(20)-[cis]-pregnen-2l-oate melting at 206 to 209 degrees centigrade having an [u] of plus 37 degrees PREPARATION .2

To a solution of 1.73 grams of 3-ethylene ketal of methyl 3,1 1 diketo 50:,6oc oxido-l7(20)-icisl-;:regnen- .21-ate in sixteen milliliters of methylene chloride was added six milliliters of 48 percent hydrofluoric acid. The heterogeneous mixture was stirred for two hours, made slightly basic with 300 milliliters of five percent sodium bicarbonate solution, and extracted with methylene chloride. The extract was washed, dried and evaporated to dryness to give 1.62 grams of crude solid. Chromatography gave two fractions: A, 481 milligrarns eluted with methylene chloride plus five percent acetone and B, 921 milligrams eluted with methylene chloride plus ten and twenty percent acetonei Crystallization of fraction' A from acetone-Skellysolve B hexanes gave 390 milligrams of methyl 3,11-diketo-a-hydroxy-6 9-fiuoro- 17(20)-allopregnen-21-oate, melting point 254 to 260 degrees centigrade. An'analytical sample melted at 260 to 263'degrees centigrade.

Analysis.Calculated for C H O F: F, 4.84. Found: F, 4.47. i

PREPARATION 3 Methyl 3,11 diketo-5a-hydroxy-6B-fluor0-17(20)- allopregnen-Zl-oate S-ethylene 'ketal pregnen-Zl-oate 3-ethylene ketal, melting point 170 to 173 degrees centigrade.

PREPARATION 4 To a solution of 1.96 grams of methyl 3,11-dlk6l0-5whydroxy 6 3 fiuoro 17(20) allopregnen 21 oate 3-ethylene ketal in 850 milliliters of anhydrous ether was added 3.7 grams of lithium aluminum hydride and the mixture was stirred for a period of one hour. 200 milliliters of water was added slowly and the ether phase separated. The aqueous phase was extracted with ethyl acetate and the extracts added to the ether phase. The combined ether-ethyl acetate solution was washed with water, dried and evaporated to dryness under reduced pressure. The crude solid residue was crystallized from acetone-Skellysolve B hexanes to give 1.30 grams of 504,115,21 trihydroxy 65 fluoro 17(20) allopregnen-3-oue 3-ethylene ketal, melting point 197 to 205 degrees centigrade. An additional 226 milligrams was obtained from the mother liquor, melting point 175 to 185 degrees centigrade.

PREPARATION 5 .5 0a,] 1 fi-dihydroxy-dfi-fluoro-ZI -acetoxy-1 7 (20 allopregnenJ-one 3-ethyiene ketal The'acetate was prepared by allowing 0.87 gram of 504,113,21- trihydroxy 6,8 fluoro 17(20) allopregnen-3-one 3-ethylene 'ketal to stand overnight in ten milliliters of acetic anhydride and ten milliliters of pyridine. The'solution was then poured into ice water to give 0.92 gram of 5 x,1lfi-dilaydroxy-6fi-fiuoro-2l-acetoxyl7(20)-allopregnen-3-one 3-ethylene ketal, melting point to degrees centigrade, which on recrystallization from acetone-Skellysolve B hexanes gave 0.77 gram, melting point 149 to 153 degrees centigrade.

PREPARATION 6 To a solution of 0.77 gram of 5a,11B-dihydroxy-6B- fiuoro 21 acetoxy 17(20) allopregnen 3 one 3- ethylene ketal in 35 milliliters of tertiary butyl alcohol was added one milliliter of pyridine, 1.9 milliliters of 1.95 molar tertiary butyl alcohol solution of N-methylmorpholine oxide peroxide, and 13.1 milligrams of osmium tetroxide (9.1 milliliters of tertiary butyl alcohol solution containing 1.44 milligrams OsO per milliliter). The solution was stirred for a period of 2.5 hours, fifteen milliliters of five percent sodium hydrosulfite added, stirred for an additional ten minutes, 0.7 gram of finely ground synthetic magnesium silicate added, stirred for a period of twenty minutes more and filtered. The filtrate was taken to dryness under reduced pressure (below fifty degrees centigrade) and the residue dissolved in methylene chloride, washed with water, dried and evaporated to dryness. This residue was crystallized from acetone-Skellysolve B hexanes to give 047 gram of 50:, 11,13,170 trihydroxy 65 fluoro 21 acetoxyallopregnane-3,20-dione 3-et-hylene ketal, melting point 220 to 228 degrees Centigrade.

PREPARATION 7 5a,11fi,17a-trihydr0xy 7 6B fluoro 21 acetoxyallopregmane-3,20-di0ne PREPARATION 8 A solution of 100 milligrams of 5a,11fi,17a-trihydroxy- 6fi-fluoro-21B-acetoxyallopregnane-3,20-dione in 4.9 milliliters of acetic acid and 0.1 milliliter of water was refluxed for a period of one hour, cooled, diluted with fifty milliliters of water and evaporated to dryness under reduced pressure. The residue was chromatographed over Plorisil (synthetic magnesium silicate) to give one fraction (77 milligrams) eluted with methylene chloride plus ten percent acetone. Crystallization from acetone- Sl-zellysolve B hexanes gave 38 milligrams of (SQ-fluoro- 11,8,17a-dihydroxy-21-acetoxy-4-pregnene-3,20-dione (6,3- iluorohydrocortisone acetate), melting point 210 to 218 egrees centigrade. Infrared data and ultraviolet data were found to he in agreement with the structure.

PREPARATION 9 Isomerization of 6 3- 10 6OL-flLlOi'OhydI'0COIZiSOFZE acetate A solution of 0.132 gram of 6/3-flu0rohydrocortisone acetate in twelve milliliters of chloroform and 0.1 milliliter of-absolute alcohol was cooled to minus ten degrees centigrade in an ice-salt bath and a stream of anhydrous hydrochloric acid was gently bubbled through the solution -for 2.5 hours While the temperature was maintained between minus'five and minus fifteen degrees centigrade.

' with water.

PREPARATION 1O 6a-fluoro-11/i,]7a,2I-trihydr0xy 4 pregnene-3,20-dione (6 ocfl uorohydrocortisone A solution of 1.1 grams of 6a-fluorohydrocortisone acetate, 1.0 gram of potassium bicarbonate, 100 milliliters of methanol and fifteen milliliters of water was purged with nitrogen and stirred at 25 degrees centigrade for four hours. The solution was then neutralized by addition of acetic acid and the methanol was removed by distillation under reduced pressure. The residue was extracted with 100 milliliters of methylene dichloride and the extract, after drying over sodium sulfate, was chromatographed over a column of eighty grams of synthetic magnesium silicate. The product fraction was eluted with Skellysolve B hexanes plus twenty and thirty percent acetone and gave 770 milligrams of 6a-fluorohydrocortisone which melted at 192 to 195 degrees centigrade after crystallization from ethyl acetate-Skellysolve B hexanes. An analytical sample melted at 192 to 201 degrees centigrade and had a rotation of [:1 plus 127 degrees (chloroform).

Analysis.-Calcd. for C H O F: C, 66.29; H, 7.68; F, 4.99. Found: C, 66.28; H, 7.65; F, 4.43.

PREPARATION 1 l 1-dehydr0-6a-flu0roh3 drocortisone Five 100-milliliter portions of a medium in 250-milliliter Erlenmeyer flasks, containing one percent glucose, two percent corn steep liquor (sixty percent solids) and tap water, were adjusted to a pH 4.9. This medium was sterilized for 45 minutes at fifteen pounds per square inch pressure and inoculated with a one to two day growth of Septomyxa afiinis, A. T. C. C. 6737. The Erlenmeyer flask was shaken at room temperature (about 26 to 28 degrees centigrade) for a period of three days. At the end of this period this 500-milliliter volume was used as an inoculum for ten liters of the same glucose-corn steep liquor medium which, in addition, contained ten milliliters of an antifoam compound (a mixture of lard oil and octadecanol). The fermentor was placed into the water-bath, adjusted to 28 degrees centigrade and the content stirred (300 R. P. M.) and aerated (0.3 liter air per minute to five liters of beer). After 24 hours of incubation, when a good growth had been developed, five grams of 6a-fluorohydrocortisone acetate plus one-half gram of 3-ketobisnor-4-cholen-22-al, dissolved in 25 milliliters of dimethylforrnamide was added and the incuba-- tion carried out at the same temperature (28 degrees centigrade) and aeration for a period of 72 hours (final pH 8.3). The mycelium was filtered off and washed The wash water was combined with the filtrate and the whole was extracted with three two-liter portions of a mixture of methylenechloride-ethyl acetate (3:1). Removal of the solvent by evaporation gave 5.25 grams of crude solid which was triturated twice with four milliliters of methylene chloride to give 2.4 grams of 1-dehydro-6a-fluorohydrocortisone of melting point 198 to 203 degrees centigrade. An analytical sample, recrystallized from acetone, melted at 202 to '204 degrees centigrade. Analysis gave [ot] plus 73 degrees (dioxane) and the following Analysis.-Calcd. for C H O F: C, 66.65; H, 7.10; F, 5.02. Found: C, 66.69; H, 7.19; F, 5.49.

PREPARATION 12 6a-fluor0-17u,21-dihydr0xy 4,9(11) pregnadiene-3,20-

dione 21 -acetate To a solution of one gram of 6u-fluorohydrocortisone acetate in ten milliliters of pyridine was added 0.4 gram of N-bromoacetamide. The mixture was allowed to stand under nitrogen for twenty minutes, at which time it was cooled to five degrees centigrade. While stirring anhydrous sulfur dioxide was passed over the surface until the solution gave no color change with acidified starch-iodide paper. The temperature of the reaction mixture was not allowed to go above twenty degrees centigrade during the sulfur dioxide addition. The mixture was then allowed to stand for five minutes and was poured into milliliters of ice-water, resulting in precipitation of 915 milligrams of crude solid, melting point to 202 degrees centigrade. Crystallization from acetone gave 511 milligrams of 6ot-fluoro17a,21-dihydroxy-4,9(11)- pregnadiene-3,20-dione 21-acetate, melting point 214 to 218 degrees centigrade. An analytical sample melted at 220 to 227 degrees centigrade. Analysis gave [al plus 73 degrees (acetone) and the following:

Analysis.Calcd. for C H O C, 68.30; H, 7.23; F, 4.70. Found: C, 68.77; H, 7.57; F, 4.77.

PREPARATION 13 6a-flu0r0-9abrom0 11,8,17oc,21 trihydr0xy-4-pregnene- 3,20-di0ne ZI-acetate (6a-fluoro-9a-bromo-hydrocortisone acetate) i To a solution of 420 milligrams of 6u-flll0I'O-17a,2ldihydroxy-4,9(11')-pregnadiene-3 ,20-dione 21-acetate in 6.5 milliliters of methylene chloride was added 12.5 milliliters of tertiary butyl alcohol, a solution of 1.0 milliliter of 72 percent perchloric acid in 75 milliliters of water, and a solution of 182 milligrams of N-bromoacetamide in 3.2 milliliters of tertiary butyl alcohol. After stirring for fifteen minutes, a solution of 182 milligrams of sodium sulfite in ten milliliters of water was added and the mixture concentrated under reduced pressure at sixty degrees centigrade until crystallization occurred. After cooling in an ice bath, thirty milliliters of water was added with stirring. The crystalline product was filtered, washed with water and dried, giving a yield of 480 milligrams of essentially pure Got-fiLlOIO-9a-bIOII10-1 1[3,17a,21- trihydroxy 4 pregnene-3,20-dione 2l-acetate, melting point 163 to 166 degrees centigrade (with decomposition). The product can be used in the succeeding example without further purification.

PREPARATION 14 6 Ot-fl tram-9,8,1 1B-0xid0-17ot,21-dihydr0xy-4-pregnenm 3,20-di0ne 2] -acetate A mixture of 2.816 grams of 6a-fluoro-9a-bromv- 1 1 ,8,l7a,2l-trihydroxy-4-pregnene-3,20-dione 21-acetats (6a-fluoro-9tx-bromohydrocortisone acetate), 2.816 grams of potassium acetate, and ninety milliliters of acetone was stirred and heated at reflux temperature for eighteen hours. The reaction mixture was then concentrated to about one-half the original volume and cooled in an ice bath. Addition of 250 milliliters of water gave 2.264 grams of 6oc-fi110IO-9fi,llB-OXidO-170:,21-dihYdI0XY-4-P1'65- nene-3,20-dione 21-acetate, melting point to 200 degrees centigrade with decomposition. The analytical sample, recrystallized from acetone, melted at 197 to 200 degrees centigrade. Analysis gave [111 plus 28 degrees (acetone) and the following:

Analysis.-Calcd. for C H O F: C, 65.70; H, 6.95; F, 4.52. Found: C, 65.76; H, 7.03; F, 4.24.

PREPARATION 15 6a,9u-diflu0r0 11B,I7a,21 trihydr0xy-4-pregnene-3,20- dione ZZ-acetate (6:,9a-difluorohydrocortisone acetate) To 3.41 grams of liquid hydrogen fluoride cooled in a poured slowly into a stirred mixture of 300 milliliters ice-water, fifty milliliters of methylene chloride, and twenty grams of sodium bicarbonate. The mixture was stirred for a few minutes, the methylene chloride layer was separated and the water phase extracted with two fifty-milliliter portions of fresh methylene chloride. The combined methylene chloride solutions were washed with water and dried. Attempts to crystallize the product by addition of Skellysolve B hexanes gave only oil. The oil was again dissolved by addition of methylene chloride and chromatographed over synthetic magnesium silicate. One long fraction of 1.496 grams of crystalline product came down from the column, which, according to papergram analysis, was a mixture of several components.

The whole column fraction was acetylated overnight with ten milliliters of acetic anhydride in ten milliliters of pyridine. The acetylation mixture was poured into ice-water andextracted with methylene chloride. The

extract was washed with dilute acid, dilute base, 'water,

PREPARATION l6 Nitrogen was bubbled through a solution of 0.33 gram of 6u,9a-difluoro-115,17a,21-trihydroxy-4-pregnene-3 ,20- dione 21-acetate (6a,9a-difiuorohydrocortisone acetate) in 33 milliliters of methanol for fifteen minutes. To this was added a solution of 0.33 gram of potassium bicarbonate in four milliliters of water, likewise treated with nitrogen. After stirring under nitrogen for five hours, the base was neutralized by the addition of 2.5 milliliters of five percent hydrochloric acid. The mixture was then concentrated under reduced pressure at fifty degrees centigrade to about five milliliters. The residue was taken up'in ethyl acetate, washed with water, dried and evaporated to dryness. Crystallization of the residue from acetone-Skellysolve B hexanes yielded 0.27 gram of 611,904- difiuoro 11B,17oc,21 trihydroxy 4-pregnene-3,20-dione (6a,9et-difiuorohydrocortisone), melting point 210 to 218 degrees centigrade. to 217 degrees centigrade. Analysis gave [al plus 1 15 degrees (acetone) and the following:

Analysis.Calcd. for C21H2805F2: C, H, F, 9.54. Found: C, 63.60; H, 7.39; F, 8.48.

PREPARATION 17 6a-fluoro 1113,17 dihydroxy-21fi-acetoxy-L4-pregnadiene-3,20-dine (1 -dehydro 6a fluorohydrocortisone acetate) A solution of two grams of 1-dehydro-6a-flu0rohydrocortisone in ten milliliters of pyridine and ten milliliters of acetic anhydride was allowed to stand at room temperature for seventeen hours, and was then poured into a mixture of ice and water. The resulting crystalline product was isolated by filtration, washed with water and dried. The yield of 1-dehydro-6a-fiuorohydrocortisone acetate was 1.32 grams of melting point 232 to 237 de- An analytical sample melted at 214' grees centigrade. An analytical sample melted at 238 to 242 degrees centigrade. Analysis gave [al plus 102 degrees (acetone) and the following:

Analysis.-Calcd. for C23H2905F1 C, H, F, 4.52. Found: C, 65.58; H, 7.16; F, 4.39.

PREPARATION 18 6a fluoro 17a hydroxy 21 acetoxy 1,4,9(11)- pregnatriene-3,20-dione To a solution of 1.05 grams of.1-dehydro-6a-fluorohydrocortisone acetate in ten milliliters of pyridine was added 0.517 gram of N-bromoacetamide. The mixture was allowed to stand under nitrogen for fifteen minutes, at which'time it was cooled to five. degrees centigrade. While stirring, sulfur dioxide was passed over the'surface until the solution gave no color change with acidified starch-iodide paper. The temperature of the reaction mixture was not allowed to go above twenty degrees centigrade during the sulfur dioxide addition. The mixture was, then poured into milliliters of ice-water, resulting in precipitation of 977 milligrams of 6ot-fluoro- 1701 hydroxy 21 acetoxy 1,4,9(11) pregnatriene- 3,20-dione, melting point 186 to 196 degrees centrigrade (with decomposition). An analytical sample melted at 21.3 to 216 degrees centigrade (with decomposition). Aanalysis gave [011 plus 34 degrees (acetone) and the following: I

Analysis-Calm. for 0 11 0 1 C, 68.64; H, 6.76; F, 4.72. Found: C, 68.85;}1, 6.86; F,- 4.72.

PREPARATION 19 600 -.flu0r0 9a bronto 11B, dihydroxy 21 ace- ;t0xy 1,4 preglmdiena- 3,20 dione (1 dehydro 60a fiuoro 906- bromohydrocoi'tisone acetate) Toa solution of 1.27 grams of 601- fluoro 17m hydroxy 21 acetoxy 1,4,9(l1) pregnatriene 3,20- dione in 19.5 milliliters of methylene chloride was added 38 milliliters of tertiary butyl alcohol, a solution of three milliliters of 72 percent perchloric acid in 22.5 milliliters of water, and a solution of 0.55 gram of N-bromoacetamide in 9.6 milliliters of tertiary butyl alcohol. After stirring for fifteen minutes, a solution of 0.55 gram of sodium sulfite in thirty milliliters of water was added and the mixture concentrated under reduced pressure at sixty degrees centigrade until crystallization occurred.

After cooling in an ice bath, 100 milliliters of water was added with stirring. 'On filtering the crystalline product, followed by washing with water and drying, a yield of 1.59 grams of essentially pure 6a-fluoro-9abromo 11 8,17 dihydroxy 21 acetoxy 1,4-preg- PREPARATION 20 6a fluoro 9,8,115 oxido 17a hydroxy ZI-acetoxy- 1,4-pregnadiene 3,20-di0ne A mixture of 1.749 grams of 6a fiuoro 9a bromo 115,170; dihydroxy 21 4 acetoxy 1,4 pregnadiene- 3,20-dione (1-dehydro-6ca-fluoro-9a-bromohydrocortisone acetate) 1.749 grams of potassium acetate, and fifty milliliters of acetone was stirred and heated at reflux temperature for eighteen hours. The reaction mixture was then concentrated to about one-half the original volume cooled and poured into 300 miliiliters of water to give 1.303 grams of 6m fiuoro 913,116 oxido 17a-hYdIOXY- 21' acetoxy 1,4 pregnadiene 3,20 dione, melting point 234 to 238 degrees centigrade (with decomposition). An analytical sample, recrystallized from acetone, melted at 257 to 260 degrees centigrade. Analysis gave [061]) plus seventy degrees (acetone) and the following: Analysis.Calcd. for C H O F: C, 66.01; H, 6.50; F, 4.54. Found: C, 65.73; H, 6.58; F, 3.87.

9 1 1 PREPARATION 21 To 5.2 grams of liquid hydrogen fluoride cooled in a Dry-Ice bath, was added, portion-wise, a slurry of 2.276 grams of 60a fiuoroj- 9,63,11,43 oxido 17a hydroxy- 2l-acetoxy-1,4-pregnadiene-3,ZO-dione in nine grams of tetrahydrofuran (distilled over NaOH) and 28 milliliters of methylene chloride which had similarly been cooled in a Dry-Ice bath. The steroid dissolved completely. After standing at zero to five degrees centigrade for seventeen hours, the reaction mixture was poured slowly into a stirred mixture of 500 milliliters of water and 25 grams of sodium bicarbonate. The mixture was stirred for a few minutes, and the product was extracted with three 100-milliliter portions of methylene chloride. The methylene chloride solutions were washed with water, dried, and chromatographed over synthetic magnesium silicate. Thefraction eluted from the column with fifteen and'twenty percent acetone in Skellysolve B hexanes was recrystallized from ethyl acetate-'Skellysolve B hexanes and gave 1.342 grams of 1 dehydro 60,9tx-difl11010- hydrocortisone acetate, melting point 238 to 242 degrees Centigrade. An analytical sample melted at 239 to 242 degrees centigrade. Analysis gave [041 plus 91 degrees (acetone) and the following:

Ar'zalysis. Calcd. for C H O F C, 63.00; H, 6.44; F, 8.67. Found: C 63.23; H, 6.82; F, 8.14.

PREPARATION 22 Nitrogen was bubbled through a solution of 1.4 grams of 6(X,9l1 difluoro 1119,1712 dihydroxy 21 acetoxy- 1,4 pregnadiene 3,20 dione (1 dehydro 601,901- difiuorohydrocortisone acetate) in 140 milliliters of methanol for fifteen minutes. To this was added a solution of 1.4 grams of potassium bicarbonate in 17.5 milliliters of water likewise treated with nitrogen. After stirring under nitrogen for five hours, the base was neutralized by the addition of 1.5 milliliters of acetic acid in forty milliliters of water. The mixture was then concentrated under reduced pressure at 55 degrees centigrade until crystallization started. The slurry was then cooled in an ice bath, diluted with 100 milliliters of water, and filtered to give 0.892 gram of 611,906 difluoro l1fl,17oc,21-1I1 hydroxy 1,4 pregnadiene 3,20 dione (1 dehydro- 6a,9 x-difluorohydrocortisone), melting point 232 to 242 degrees centigrade (with decomposition). An analytical sample melted at 250 to 257 degrees Centigrade (with decomposition). Analysis gave [061]) plus 84 degrees (acetone) and the following:

Analysis.Calcd. for c r-1, 1%; c, 63.62; H, 6.61;

F, 9.59. Found: C, 62.26; H, 7.10; F, 9.41.

PREPARATION 23 To a solution of 770 milligrams of crude Got-fluoro- 1 1,8,1701,21-trihydroxy-4-pregnene-3,20-dione (6a-fiuorohydrocortisone) in ten milliliters of pyridine previously cooled to zero to five degrees centigrade there was added 0.7 milliliters of methanesulfonyl chloride. The reaction mixture was stirred in an icewater bath for four hours. Dilution with 100 milliliters of five percent hydrochloric acid precipitated the crystalline mesylate. The product, after filtration, weighed 900 milligrams and melted at 189 to 192 degrees (with decomposition). Infrared analysis in mineral oil mull showed adsorptions as follows: 3560, 3420 centimeters (OH); 1725 centimeters- (20- ketone); 1655 centimeterr (A -3-ketone); 1640, 1617 centimeters (C=C); 1350,1200, 1170 centimeters- (OSO2).

PREPARATION 24 A mixture of 200 milligrams of crude mesylate and 200 milligrams of sodium iodide in ten milliliters of acetone was heated under reflux for fifteen minutes. The solvent was distilled under vacuum, and the resulting 6oc-flll0l0- 1 f3,1'7a-dihydroxy-4-pregnene 3,20 dione was not purified but was employed in the crude state in the succeeding step of Preparation 25.

PREPARATION 25 6ot-flltOiO-J 113,1 7u-dihydroxy-4-pregnene-3,20-di0ne The crude product from Preparation 24 above was dissolved in four milliliters of acetic acid. After stirring continuously at room temperature for thirty minutes, sulficient five percent sodium thiosulfate solution was added to discharge the iodine color. The resulting colorless solution was poured into milliliters of water containing five grams of potassium bicarbonate. On filtering the resulting crystals, milligrams of crystalline product melting at to degrees Centigrade was obtained. Several recrystallizations from ethyl acetate yielded a product melting at 219 to 222 degrees centigrade. This roduct was identified as 6a-fluoro-11B,17a-dihydroxy-4- pregnene-3,20-dione, with infrared adsorptions as follows: 3590, 3380 centimeters- (OH); 1697 centimeters (20-ketone); 1665- centimeters (conjugated ketone); 1623 centimeters" (C=C). Analysis was as follows:

Analysis.Calcd. for C I-1 F0 1 C, 69.20; H, 8.02; F, 5.21. Found: C, 69.59; H, 8.09; F, 3.97.

. PREPARATION 26 6a-flu01'0-11/3J 7ot-dihydr0xy-1,4-prenadiene-3,20-di0ne 300 milligrams of 66t-fiuoro-11B,17a,21-trihydroxy-1,4- pregnadiene-3,20-dione was dissolved in three milliliters of pyridine and cooled to zero to five degrees centigrade. One tenth of one milliliter of methanesulfonyl chloride was added and the solution maintained at zero to five degrees centigrade for sixteen hours. When poured into a solution of three milliliters of concentrated hydrochloric acid in fifty milliliters of water, crystalline 6ot-fiuoro-11fi, 17,21-trihydroxy-1,4-pregnadiene-3 ,20-dione 21-methanesulfonate precipitated. The yield of product was 310 milligrams and melted at 200 to 202 degrees centigrade with decomposition. Infrared absorption in a mineral oil mull was as follows: 3570, 3370 centimeters- (OH); 1727 centimeters (20-ketone); 1665 centimeters- (conjugated ketone); 1623, 1601 centimeters- (A 1360, 1342, 1172 centimeters- (080 A mixture of 100 milligrams of 6a-fluoro-l 1,8,17a,21- trihydroxy-l,4-pregnadiene 3,20 dione 21-methanesulfonate and 100 milligrams of sodium iodide in five milliliters of acetone was refluxed for fifteen minutes. The solvent was removed by vacuum distillation. The crude 6a-fluoro-21-iodo-11p,17a-dihydroxy 1,4 pregnadiene-3, 20-dione was not isolated but dissolved in two milliliters of acetic acid. After fifteen minutes sufiicient aqueous sodium thiosulfate was added to discharge the iodine 1 3 PREPARATION 27 To a solution of 500 milligrams of 6a,9a-difluoro-11/3, 17m,21-trihydroxy-4-pregnene-3,20-dione in six milliliters of pyridine previously cooled to zero to five degrees centigrade was added 0.55 milliliter of methanesulfonyl chloride. The reaction mixture was stirred at six degrees centigrade for sixteen hours and then poured into 100 milliliters of cold five percent hydrochloric acid solution. The resulting solid mesylate, 6a,9ot-difluoro-11fl,17a,21 trihydroxy-4-pregnene 3,20 dione ZI-methanesulfonate, after being filtered, washed with water and dried, weighed 620 milligrams.

PREPARATION 28 6a,9a difluoro 115,17a dihydroxy 21 iodo 4 pregnene-3,20-a'i0ne and 6a,9ot-difluor0-11fi,17a-dihydrxy-4-pregnene-3,20-di0ne To a solution of 370 milligrams of crude 6a,9a-difluoro- 11,6,17u,21-trihydroxy-4-pregnene-3,20-dione ZI-methanesulfonate in ten milliliters of acetone was added a solution of 370 milligrams of sodium iodide in four milliliters of acetone. The mixture was stirred at room temperature for twenty minutes and at reflux for fifteen minutes and was then evaporated to dryness at reduced pressure. The residue containing 60,9zx-diflllOIO-l113,17oc-dlhYdlOXY- 2l-iodo-4-pregnene-3,20-dione was dissolved in five milliliters of acetic acid and stirred for 45 minutes. A solution of 600 milligrams of sodium thiosulfate in 7.2 milliliters of water was added to the acetic acid solution, followed by seventy milliliters of ice and water. The resulting solid, isolated by filtration, weighed 185 milligrams and was chromatographed over synthetic magnesium silicate and crystallized from acetone-Skellysolve B hexanes mixture to give 137 milligrams of 6u,9a-difluoro-11B,17udihydroxy-4-pregnene-3,20-dione, melting at 220 to 240 degrees centigrade. An additional 59 milligrams of product melting at 200 to 220 degrees centigrade was obtained by extracting the water phase from the above precipitation step with methylene chloride, followed by chromatographing and crystallizing as above. The total crystalline material was recrystallized several times from ethyl acetate to give an analytical sample which melted at 253 to 256 degrees centigrade. Analysis gave [aJ plus 109 degrees (acetone) and the following:

Analysis.Calcd. for C l-1 0 1 C, 65.95; H, 7.38; F, 9.94. Found: C, 66.26; H, 7.49; F, 9.10.

PREPARATION 29 6a,9a-difluor0-11l3,17a,21-trihydr0xy-1,4-pregnadiene- 3,20-di0ne 21 -methanesulf0nate To a solution of 0.925 gram of 6a,9tx-difiuoro-11fl,17a, 21-trihydroxy-l,4-pregnadiene-3,20-dione in ten milliliters of pyridine previously cooled to zero to five degrees centigrade was added 0.9 milliliter of methanesulfonyl chloride. The reaction mixture was stirred at zero to five degrees centigrade for seventeen hours, and was then poured into 100 milliliters of cold five percent hydrochloric acid to precipitate the solid mesylate. The product, after filtration, weighed 0.832 gram and had a melting point of 157 degrees centigrade (with decomposition).

PREPARATION 3 O 6ot,9ot difluoro 115,17u dihydroxy 21 iodo 1,4 pregnadiene-3,20-di0ne and 60,9ct-diflbl07'0-11fl,17oz-di hydroxy-l,4-pregnadiene-3,20-di0ne To a solution of 332 milligrams of 6u,9a-difiuoro-11/8,- 17a,21-trihydroxy-1,4-pregnadiene-3,20-dione 21-methanesulfonate in eight milliliters of acetone was added a solution of 332 milligrams of sodium iodide in eight milliliters of acetone. The mixture was stirred at reflux for fifteen minutes and was then evaporated to dryness with a stream of nitrogen. The residue containing 60,9a-difiuoro l1fi,17u dihydroxy 21 iodo 1,4 pregnadiene-3,20-dione was dissolved in eight milliliters of acetic acid and stirred for 45 minutes. A solution of 400 milligrams of sodium thiosulfate in eight milliliters of water was added to the acetic acid solution, then milliliters of ice and water was added. The resulting solid, isolated by filtration, weighed 272 milligrams, and was chromatographed over synthetic magnesium silicate, then crystallized from acetone to give 127 milligrams of 611,911- difluoro 11,8,1'7a dihydroxy 1,4 pregnadiene 3,20 dione, melting at 288 to 295 degrees centigrade. The analytical sample melted at 296 to 300 degrees centigrade. Analysis gave [al plus 58 degrees (pyridine) and the fol lowing:

Anaiysis.-Calcd. for C H O F C, 66.30; H, 6.89; F, 9.99. Found: C, 65.88; H, 6.76; F, 9.39.

One hundred milliliters of two percent cornsteep liquor of sixty percent solids was adjusted to pH of 6.8 to 7.4 with sodium hydroxide and was sterilized at fifteen pounds pressure for thirty minutes. To this was added a similarly sterile solution of two grams of Cerelose (technical grade of dextrose) in the four milliliters of water. This sterile medium was inoculated with a suspension of spores and mycelium of Streptomyces roseochromogenus (Waksman Collection No. 3689) and was agitated on a rotatory shaker for a period of 24 hours by which time a good growth of the organism had taken place. To this 24 hour culture, twenty milligrams of 6ot-fiuoro-1 1,8,17adihydroxy-4-pregnene-3,20-dione dissolved in 0.2 milliliter of dimethylformamide was added. Incubation of the steroid with the microorganism was maintained (with agitation) for five days, at which time the pH was 8.6. The fermentation broth was then separated into the mycelium and the beer by centrifugation. The mycelium was extracted first with two 25 milliliter portions of acetone and then with four successive 25 milliliter portions of methyl isopropyl ketone. The beer was extracted with four successive 25 milliliter portions of methyl isopropyl ketone. All of the extracts were combined, washed with two percent aqueous sodium bicarbonate solution and with water, dried with anhydrous sodium sulfate, and evaporated to dryness. The residue, which on paper chromatogram analysis showed the presence of 6OL-flllOIO-11[3,16zx,- 17a-trihydroxy-4-pregnene-3,ZO-dione, was purified by chromatography over synthetic magnesium silicate (Florisil) and crystallization from acetone to give 6t-.-fiuoro- 11B,16oz,17ot-trihydroxy-4-pregnene-3,20-dione.

Following the procedure of Example 1 above, but substituting 6a,9a-difiuoro-11B,17a-dihydroxy-4-pregnene-3,- 20-dione, 60L-flLlOI0-1 l 5,17a-dihydroxy-1,4-pregnadiene-3,- ZO-dione, or 6u,9oc-difluoro-11(3,17a-dihydroxy-1,4-pregna-- diene-3,20-dione as starting material therein, is productive of 60:,9ot-diflll01O-11f ,l60t,170t-tIlhydI'OXy-4 p1'gfl(5!l6 3,20-dione, 6a-fiuoro-1 l 6,1611, 1 7ot-trihydroxy-1,4-pregna diene-3,20-dione, and 6a,9oz-difluoro-l1,8,16a,17a-trihydroxy-l,4-pregnadiene-3,20-dione, respectively.

EXAMPLE 2 A solution of 1.2 grams of 6a-fluoro-11,B,16a,17ot-trihydroxy-4-pregnene-3,20-dione (from Example 1) in twenty milliliters of pyridine and twenty milliliters of acetic anhydride was allowed to stand at room temperature (about 25 degrees centigrade) for eighteen hours and was then poured into 200 milliliters of ice-water. The resulting mixture was extracted with methylene chloride and the extract was washed with dilute hydrochloric acid, dilute sodium bicarbonate, and water. After drying the solution with anhydrous sodium sulfate, the solvent was removed by evaporation and the residue was purified by chromatography over synthetic magnesium silicate (Florisil) and crystallization from acetone to give 6a-fluoro- 1lfl,16a,17u-trihydroxy-4-pregnene-3,ZO-dione 16-acetate.

Following the procedure of Example 2 above but substituting 60:,90c-difl11010-1lfi,l6oz,170c-trlhyd10Xy 4 pregnene-3,20-dione, 6a-fiuoro llp,l6a,l7u trihydroxy-1,4- pregnadiene-3,20-dione or 60:,9a-diflllOIO-11B,160t,17ot-tri hydroxy-1,4-pregnadiene-3,20-dione as starting material therein is productive of 6u,9a-difluoro-11fi,16u,17o-trihydroxy-4-pregnene-3,20-dione 16-acetate, 6a-fiuoro-1lfi, 16oz, 17a-trihydroxy-1,4-pregnadiene-3,20-dione 16-acetate and 6a,9a difiuoro-l 15,1611, 17a-trihydroxy-1,4-pregnadicue-3,20-dione 16-acetate, respectively.

Similarly, acylation of 6u-fluoro-11B,16u,17a-trihydroxy-4-pregnene-3,20-dione with the appropriate acid anhydride or acid chloride is productive of still other 16-acylates such as, for example, 6oc-fluOr0-11}3,16a,17atrihydroxy-4-pregnene-3,ZO-dione 16-propionate, 6oc-fllloro-l1,8,16a,17a-trihydroxy-4-pregnene 3,20 dione 16- butyrate, 6a-fiuoro-1 15,1611,17a-trihydroxy-4-pregnene-3, -dione l6valerate, 6a-fluoro-llB,16a,l7a-trihydroxy-4- pregnene-3,20-dione l6-hexanoate, 6oc-fil10r0-11/3,16a,17utrihydroxy-4-pregnene-3,20-dione l6-laurate, 6oc-flll0f0- 1 15,1601, l7a-trihydroxy-4-pregnene-3,20-dione 16-trimethylacetate, 6u,-fluoro-llfl,l6u,l7a-trihydroxy-4-pregnene- 3,20-dione 16-isobutyrate, 6u-fluoro 1lfl,160t,17a trihydroxy-4-pregnene-3,ZO-dione 16-is0valerate, 6a-fltlOI0-1lfi, 16oz,17a-trihydroxy-4-pregnene-3,20-dione 16-cyc1ohexane carboxylate, 6a-fluoro-1 113, 16a,17u-trihydroxy-4-pregnene- 3,20-dione l6-benzoate, 6tx-fluoro-1 1,8, 1 6oz, l7u-trihydroxy- 4-pregnene-3,20-dione 16-phenylacetate, 6a-flUOIO-11j3, 16a,17a-trihydroxy-4-pregnene-3,20-dione 16-(B-phenyl)- propionate, 6a-fluoro-l 1 3,16, l7a-trihydroxy-4-pregnene- 3,20-dione 16-(o-, m-, p-toluate), 60c-fl11010-11fi,16ot,17ot-trihydroxy-4-pregnene-3,ZO-dione l6-hemisuccinate, 6a-fi1l- OIO11fi,16a,17ot trihydroxy-4-pregnene 3,20 dione l6- hemiadipate, 6a-fluoro 11,8,16u, 17a trihydroxy-4-pregnene-3,20-dione l6-acrylate, 6oc-fll10rO-l1fi,16oc,l7u-trihy droxy-4-pregnene-3,ZO-dione 16-undecylenate, 6a-fiuoro- 11,6,16a,17a-trihydroxy-4-pregnene-3,20-dione 16-propio-, late, 60t-fiI1OI'O-11B,16OL,170t-t1'ihYdI'OXY 4 pregnene-3,20- dione l6-cinnamate, 6a-*luoro-llB,l6a,17a-trihydroxy-4- pregnene-3,20-dione l6-maleate, Ga-flllOIO-l1fl,16oc,17a-trihydroxy-4-pregnene-3,20-dione l6-citraconate.

Similarly, acylation of 6a,9oc-difluOrO-l1/3,16oc,l7o-trihydroxyl-pregnene 3,20 dione, 6a-fluoro1 1/3,16a,17atrihydroxy-l,4-pregnadiene-3,ZO-dione, or 6a,9a-difluorol1 [3,160c,170c trihydroxy-l,4-pregnadiene-3,20-dione with the appropriate acylating agent is productive of the corresponding l6-acylates. The preferred acylates are those corresponding to the acylates described above for 60- fiuoro-l 118,160,17a-trihydroXy-4-pregnene-3,20-dione.

EXAMPLE 3 6a-fluor0-I 6a,] 7a-dihydr0xy-4-pregnsite-3,11,20-

trione 16-acetate To a solution of 0.5 gram of 6a-fluoro-11B,l6u,17ottrihydroxy-4-pregnene-3,20-dione 16-acetate in twenty milliliters of acetic acid was added a solution of 0.15 gram of chromium trioxide in one-half milliliter of water. The mixture was stirred and maintained at room temperature (about degrees centigrade) for a period of four hours. Thereafter the excess oxidant was destroyed by the addition of 0.5 milliliter of methanol and the mixture was poured into 100 milliliters-of water and extracted with methylene chloride. The extract was washed with dilute sodium bicarbonate and with water, and was dried and evaporated to dryness. The residue was crystallized from acetone to give 6a-fiuoro-l6a,l7a-dihydroxy-4pregnene-3, l 1,20-trione 16-acetate.

Following the procedure of Example 3 above but substituting 60:,9oc-dlflllOI'O-11B,16oc,l7a-trihydr0Xy 4 pregnene-3,20-dio11e l6-acetate, 60c-flu0IO-1lj3,16ot,17oz-trihycal, and R is 16 droxy-1,4-pregnadiene--2,20-dione l6-acetate, or 6a,9ozdifluOI0-l1;3,16a,l7a trihydroxy 1,4 pregnadiene-3,20- dione 16-acetate as starting material therein is productive of 6a,9ot difluoro-16a,17a-dihydroxy-4-pregnene-3,11,20- trione 16-acetate, 6a-fiuoro-l6a,17ot-dihydroxy-L4-pregnadiene-3,1l,20-trione l6-acetate, and 6a,9ot-difll10l'0- 160t,170t dihydroxy-1,4-pregnadiene 3,11,20 trione 16-- acetate, respectively.

Similarly, oxidation of other 11 fi-hydroxy 16-ester compounds of Example 2 is productive of the corresponding ll-keto l6-ester compounds. The preferred ll-keto 16-ester compounds are those wherein the acyl radical is that of an organic carboxylic acid, preferably a hydrocarbon carboxylic acid containing from one to twelve carbon atoms, inclusive.

EXAMPLE 4 6a-flu0r0-16u,1 7a-dihydroxy-4-pregnene-3,l1,20-trione A solution of 1.2 grams of 6a-fluoro-16a',17a-dihydroxy-4-pregnene-3,l1,20-trione 16-acetate, two grams of potassium bicarbonate, milliliters of methanol and fifteen milliliters of water was purged with nitrogen and stirred at 25 degrees centigrade for eight hours. The solution was then neutralized by addition of acetic acid and the methanol was removed by distillation under re duced pressure. The residue was extracted with 100 milliliters of methylene dichloride and the extract, after drying over sodium sulfate, was chromatographed over a column of eighty grams of synthetic magnesium silicate, using Skellysolve B hexanes with increasing amount of acetone for elution. The product fraction from the column was crystallized from acetone to give 6a-fiuoro- C, 1 7a-dil1ydroxy-4-pregnene-3,1 l,20-trione.

Following the procedure of Example 4, saponification of 611,9 a-difiuoro- 1 6oz, 17oc-dihydroxy-4-pregnene-3,1 1,20- trione 16-acetate, 6a-fluoro-16u,17a-dihydroxy-1,4-pregnadiene-3,11,20-trione 16-acetate and 6a,9a-difluoro-16a,- 17a-dihYd10Xy- 1 ,4-pregnadiene-3 ,1 1,20-trione 16-acetate of Example 3 is productive of the corresponding hydroxy compounds, 6a,9a-difluoro-l6a,17ot-dihydroxy 4 pregnene-3,l1,20-trione, 6oc-fiuOrO-16a,17oc dihydroxy 1,4- pregnadiene-3,11,20-trione, and 6a,9a-difiuoro-16a,17ezdihydroxy-l,4-pregnadiene-3,l 1,20-trione, respectively.

It is to be understood that the invention is not to be limited to the exact details of operation or exact compounds shown and described, as obvious modifications and equivalents will be apparent to one skilled in the art, and the invention is therefore to be limited only by the scope of the appended claims.

We claim:

1. A 16-0xygenated steroid of the formula:

C=0 ----OH wherein the 1,2-carbon atom linkage is selected from the linkages consisting of single bond and double bond linkages, Y is selected from the group consisting of hydrogen and fluorine, X is selected from the group consisting of the carbonyl radical and the ,B-hydroxymethylene radiselected from the group consisting of hydrogen and the acyl radical of a hydrocarbon carboxylic acid containing from one to twelve carbon atoms, in-.

clusive.

2. A 16-oxygenated pregnene of the formula:

(EH: o=o CH3 I... OH

wherein Y is selected from the group consisting of hydrogen and fluorine, X is selected from the group consisting of the carbonyl radical and the fi-hydroxymethylene radical, and R is selected from the group consisting of hydrogen and the acyl radical of a hydrocarbon carboxylic acid containing from one to twelve carbon atoms, inclusive.

3. 6a-fiuoro-11B,16a,17a-trihydroxy-4-pregnene 3,20- theme.

4. 6a,9oc-difiuOl'0-1 118,16a,17u-trihydroxy-4-prcgnene-3,- ZO-dione.

5. 6oz-flll0l0-16oc,17oc-dihydr0Xy 4 pregnene 3,11,20- trione.

6. 6a,9a-difluoro-16a,17a-dihydroxy-4-pregnene 3,11,- 20-trione.

7. 6a-fluoro-l1p,16a,17a-trihydroxy-4-pregnene 3,20- dione 16-acetate.

8. 6oc,9oc-difluOrO-l 15,16a,17a-trihydroxy-4-pregnene-3,- ZO-dione 16-acetate.

9. 6a-fiuoro-16a,17a-dihydroxy-4-pregnene 3,11,20- trione 16-acetate.

10. 6a,9a-difluoro- 16oz, 17a-dihydroxy-4-pregnene-3,1 1,- ZO-trione 16-acetate.

18 11. A 16-oxygenated pregnadiene of the formula:

3 ---'OH i T B wherein Y is selected from the group consisting of hydrogen and fluorine, X is selected from the group consisting of the carbonyl radical and the p-hydroxymethylene radical, and R is selected from the group consisting of hydrogen and the acyl radical of a hydrocarbon carboxylic acid containing from one to twelve carbon atoms, inclusive.

12. 6oz fiuoro 11{3,15tx,17oc trihydroxy 1,4 pregnadiene-3,20-di0ne.

l3. 6a,9oc-difiuO1'O-l 15,161,l7a-trihydroxy-1,4 pregnadiene-3,20-dione.

14. 6a-fluoro-16a,17a-dihydroxy-1,4-pregnadiene-3 ,11, 20-trione.

15. 6a,9a-difiuoro-16a,17a-clihydroxy-1,4-pregnadiene- 3,11,20-t1'ione.

16. 6a-fluoro-11B,'16a,17u-trihydroxy-l,4-pregnadiene- 3,20-dione 16-acetate.

17. 6u,9a-difluoro-1 1p,16a,17a-trihydroxy-1,4-pregna diene-3,20-dione 16-acetate.

18. 6a-fluoro-16a,17u-dihydroxy-l,4-pregnadiene-3,l1,- 20-trione 16-acetate.

19. 6a,9a-difiuoro-16a,17a-dihydroxy-1,4-pregnadiene- 3,11,20-trione 16-acetate.

No references cited. 

1. A 16-OXYGENATED STEROID OF THE FORMULA: 